Proteins, after being transcribed and translated from genome, are subjected to posttranslational modifications such as phosphorylation, which directly influence functions such as signal pathway and enzymatic activity. Since proteins change qualitatively or quantitatively with development of life, differentiation, progression of diseases, environmental changes and the like, by analyzing a set of proteins (proteome) encoded by a genome, proteins may be understood cyclopaedically, and significant knowledge on the analysis of bioinformatics, the diagnosis of diseases and the development of drugs may be obtained. Especially, phosphorylation of proteins, one type of posttranslational modification of genes, is an important step in signal transmission, activation of enzymes or the like. Consequently, identification of phosphoproteins is important to understand functions of proteins.
Up to now, gel electrophoresis has been generally used as a method for identifying proteins. Specifically, a two-dimensional gel electrophoresis method has found wide acceptance because it enables the separation of proteins in high separability. Further, to distinguish phosphoprotein from the various proteins developed on the two-dimensional gel electrophoresis, immunostaining method that uses antibody and labeling method that uses radioisotopes have been known. Specifically, a protein separated by electrophoresis is immobilized on a hydrophobic membrane, and brought into contact with an antibody, for which its antigen is the desired phosphoprotein, to form an antigen-antibody complex, which is then detected with a secondary antibody labeled with an enzyme or a radioisotope.
However, these conventional methods were problematic in that they are intricate and time-consuming, and for the methods that uses radioisotopes, special facilities are required and are dangerous.
Under these circumstances, the invention of the present application has been made, and aims to provide, upon solving the problems of the prior art, a convenient method for identifying phosphoprotein in less time.